INHIBITION OF ENZYMES 



33 



the amino acids reacted are quite different in the three pH ranges used 

 (Gundlach et al., 1959 a). Reaction at pH 5.5-6.0 leads to a chromato- 

 graphically homogeneous protein which differs from the initial ribonuclease 

 only in the carboxymethylation of a single imidazole group of histidine. At 

 pH 2.8 a single methionine and at pH 8.5 a single lysine react with iodo- 

 acetate. The rate of inactivation of ribonuclease also depends on the pH: 



Fig. 1-1. Activation and inhibition of ascites tumor 

 ribonuclease by iodoacetate and p-mercuribenzoate. 

 The enzyme activity is expressed as /^moles of TCA- 

 soluble phosphate. (From Hilz and Klempien, 1959.) 



At pH 8.5-10.5 it is slow (half-reaction in 90 min and maximal inhibition 

 65%), at pH 5.5-6.2 it is faster and more complete (maximal inhibition 

 90%), at pH 4 it is again slow, and at pH 2.8 it is more rapid. This is well 

 explained on the basis of different amino acids reacting at the different 

 pH's. The inactivation with bromoacetate behaves similarly (see accom- 

 panying tabulation) (Barnard and Stein, 1959). Only one of the four histi- 



pH 



Time for 50% inhibition (min) 



4.2 

 5.3 

 7.1 

 9.0 



140 

 14 



27 

 46 



